Enhanced antigen-antibody interactions in microfluidic immunosensors can be effected by tailoring the surface free energies of the antibody immobilized surfaces to obtain the appropriate fluid-wall interactions. We report a systematic study to evaluate the surface free energies from contact angle measurements, using the LWAB method, of different antibody (anti-BSA, anti-PSA, and anti-CRP) surfaces, each immobilized separately on to non- and nanotextured Si surfaces via a stack of functionalized layers including aminosilanes of which three different types were used. The apolar surface free energy components were independent of the physical modification in the non-functionalized and the intermediate hydrolyzed surfaces where as they depended on the nature of the surface and the chemical modifications in the subsequent functionalized stages. Surface free energies of the different antibodies immobilized with the shorter chain length aminosilane (APTES) on non-and nanotextured surfaces were in the order of anti-BSA < anti-PSA < anti-CRP. A tunability of the surface free energy up to 9.6 mJ/m(2) was achieved which is reasonably significant when compared to the surface free energy window (Delta gamma(s) = 40 mJ/m(2)) of biofunctionalized surfaces. This fundamental understanding of the surface energetics of the biofunctionalized surfaces can be utilized in modulating the surface properties to design efficient immunosensors. (C) 2013 Elsevier B.V. All rights reserved.